NaOH Yeast Colony PCR - MetabolicEngineeringGroupCBMA/MetabolicEngineeringGroupCBMA.github.io GitHub Wiki

  1. Watch this YouTube video (2 min) describing the technique.
  2. Add 20 µL 20 mM NaOH to a clean 1.5 mL Eppendorf tube.
  3. Pick a small amount of cells from your plate with a yellow tip. It is important not to take too much and no agarose (see at 52 s in the video).
  4. Add the cells to the solution and swirl to mix (see at 59 s in the video).
  5. Incubate tubes at around 95°C for ten minutes. This can be done using a plastic bucket and recently boiled water from a kettle.
  6. When the 95°C incubation is over, add 180 µL TE buffer.
  7. Vortex the tube for 30 s.
  8. Spin at max speed in a micro-centrifuge for 10-20 s.
  9. Use 5 µL of the supernatant for a 20 µL PCR reaction, do not disturb the cell debris at the bottom of the tube.
  10. A standard PCR program can be used without changes.
⚠️ **GitHub.com Fallback** ⚠️